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Engineering an Escherichia coli based in vivo mRNA manufacturing platform.
Curry, Edward; Muir, George; Qu, Jixin; Kis, Zoltán; Hulley, Martyn; Brown, Adam.
Afiliação
  • Curry E; Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, UK.
  • Muir G; Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, UK.
  • Qu J; Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, UK.
  • Kis Z; Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, UK.
  • Hulley M; Bioprocess Development, AstraZeneca, Cambridge, UK.
  • Brown A; Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, UK.
Biotechnol Bioeng ; 121(6): 1912-1926, 2024 Jun.
Article em En | MEDLINE | ID: mdl-38419526
ABSTRACT
Synthetic mRNA is currently produced in standardized in vitro transcription systems. However, this one-size-fits-all approach has associated drawbacks in supply chain shortages, high reagent costs, complex product-related impurity profiles, and limited design options for molecule-specific optimization of product yield and quality. Herein, we describe for the first time development of an in vivo mRNA manufacturing platform, utilizing an Escherichia coli cell chassis. Coordinated mRNA, DNA, cell and media engineering, primarily focussed on disrupting interactions between synthetic mRNA molecules and host cell RNA degradation machinery, increased product yields >40-fold compared to standard "unengineered" E. coli expression systems. Mechanistic dissection of cell factory performance showed that product mRNA accumulation levels approached theoretical limits, accounting for ~30% of intracellular total RNA mass, and that this was achieved via host-cell's reallocating biosynthetic capacity away from endogenous RNA and cell biomass generation activities. We demonstrate that varying sized functional mRNA molecules can be produced in this system and subsequently purified. Accordingly, this study introduces a new mRNA production technology, expanding the solution space available for mRNA manufacturing.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: RNA Mensageiro / Escherichia coli / Engenharia Metabólica Idioma: En Revista: Biotechnol Bioeng Ano de publicação: 2024 Tipo de documento: Article País de afiliação: Reino Unido

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: RNA Mensageiro / Escherichia coli / Engenharia Metabólica Idioma: En Revista: Biotechnol Bioeng Ano de publicação: 2024 Tipo de documento: Article País de afiliação: Reino Unido
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